Microbial communities perform a wide range of functions, such as microbial growth, nitrogen processing in the soil, cellulose degradation, lignin degradation, dehalogenation of hydrocarbons and aromatic hydrocarbons, decomposition of organic matter in the carbon cycle, and environmental remediation such as oxidation of hydrocarbons. Many tasks are difficult or impossible for a single microorganism to perform. These functions typically require balancing competition and interactions among multiple species. For example, bacteria form biofilms and other forms of interactions, where several types of bacteria survive through symbiotic relationships, in which nutrients and signaling molecules are exchanged. These processes may be better performed by a synthetic community of microorganisms. Furthermore, some of these processes may require microorganisms that cannot be cultured without other microorganisms present.
The vast majority of bacteria cannot be cultured and cannot be studied with traditional techniques. Under homogeneous laboratory conditions, most attempts to co-incubate multiple microbial species do not result in stable communities due to lopsided competition for nutrients among the cultured species. Attempted methods of co-incubate have potential problems: different microorganisms may be toxic to one another on contact; they may need to be kept apart with within diffusion contact; one microorganism may grow at a much faster rate than others; shared molecules such as nutrients are lost by diffusion unless the microorganisms are cultured close together.
It is known that spatial structure influences competition and interactions between microbes. Environments in which the microbes live are highly spatially heterogeneous, and it has been shown that microbes exist as isolated patches. In nature, microbial communities inhabit matrices with intricate spatial structure, for example, many species of soil bacteria stably coexist as microcolonies separated by a few hundred micrometers. Thus, spatial structure is important in microbial ecology. However, spatial structure is difficult to control in natural environments and has not been systematically experimentally varied to understand its effect on the stability of bacterial communities.
It is also known that microbial communities perform important functions that require the stable interaction of multiple species. The combined effects of competition and chemical communication dictate that some multispecies communities have required spatial structures in order to stably function. The trade-off between reduced competition and effective communication define a specific range of spatial structures for each community. Spatial structures thus may be able to stabilize microbial communities. Natural habitats provide such spatial structures and recreating some communities in the laboratory will require culturing the microorganisms in specific structures and under specific conditions.
Aspects of methods for culturing communities of interacting bacteria, are disclosed in, for example: Swenson et al., 2000, Environmental Microbiology 2: 564-571 (artificial selection of microbial ecosystems for 3-chloroaniline biodegradation); in Swenson et al., 2000, Proc. Natl. Acad. Sci. USA 97: 9110-9114 (artificial ecosystem selection); and in Williams and Lenton, 2007, Proc. Natl. Acad. Sci. USA 104: 8918-8923 (artificial selection of simulated microbial ecosystems).
Aspects of how mixed communities do not survive due to competition are disclosed in, for example: Dechesne et al., 2008, FEMS Microbiology Ecology 64: 1-8 (limited diffusive fluxes of substrate facilitate coexistence of two competing bacterial strains); in Ferrari et al., 2005, Applied and Environmental Microbiology 71: 8714-8720 (microcolony cultivation on a soil substrate membrane system, which selects for previously uncultured soil bacteria); in Treves et al., 2002, Microbial Ecology, 45: 20-28 (two-species test of the hypothesis that spatial isolation influences microbial diversity in soil); in Hassell et al., 1994, Nature 370: 290-292 (species coexistence and self-organizing spatial dynamics); in Allison, 2005, Ecology Letters 8: 626-635 (cheaters, diffusion, and nutrients constrain decomposition by microbial enzymes in spatially structured environments); in Kerr et al., 2002, Nature 418: 171-174 (local dispersal promotes biodiversity in a real-life game of rock-paper-scissors); and in Rainey and Travisano, 1998, Nature 394: 69-72 (adaptive radiation in a heterogeneous environment).
Aspects of how dependent microbial strains require close proximity are disclosed in, for example: Nunan et al., 2003, FEMS Microbiology Ecology 44: 203-215 (spatial distribution of bacterial communities and their relationship with the micro-architecture of soil); in Hansen et al., 2006, Nature 445: 533-536 (evolution of species interactions in a biofilm community); and in Nielsen et al., 2000, Environmental Microbiology 2: 59-68 (role of commensal relationships on the spatial structure of a surface-attached microbial consortium).
Aspects of how function and viability of some strains requires partner strains are disclosed in, for example, Ohno et al., 1999, Biosci. Biotechnol, Biochem, 63: 1083-1090 (establishing the independent culture of a strictly symbiotic Bacterium Symbiobacterium thermophilum from its supporting Bacillus strain); in Kaeberlein et al., 2002, Science, 296: 1127-1129 (isolating “uncultivable” microorganisms in pure culture in a simulated natural environment); in Ou and Thomas, 1994, Soil Science Soc. Am. J. 58: 1148-1153 (influence of soil organic matter and soil surface on a bacterial consortium that mineralizes fenamiphos, a pesticide); in Kato et al., 2005, Applied and Environmental Microbiology 71: 7099-7106 (stable coexistence of five bacterial strains as a cellulose-degrading community); in Price-Whelan et al., 2006, Nature Chemical Biology 2: 71-78 (rethinking ‘secondary’ metabolism: physiological roles for phenazine antibiotics); and in Cosgrove at al., 2007, Applied and Environmental Microbiology 73: 5817-5824 (different fungal communities are associated with degradation of polyester polyurethane in soil).
Aspects of how soil is spatially complex, with patchy distribution of microbes, are disclosed in, for example, Young and Crawford, 2004, Science 304: 1634-1637 (interactions and self-organization in the soil-microbe complex); and in Grundmann et al., 2001, Soil Science Soc. Am. J. 65: 1709-1716 (spatial modeling of nitrifier microhabitats in soil).
Aspects of methods for in vitro spatial culture are disclosed in, for example, Abhyankar and Beebe, 2007, Anal. Chem. 79: 4066-4073 (spatiotemporal micropatteming of cells on arbitrary substrates); in Weibel at al., 2007, Nature Reviews Microbiology 5: 209-218 (microfabrication meets microbiology); in Keymer at al. 2006, Proc. Natl. Acad. Sci. USA 103: 17290-17295 (bacterial metapopulations in nanofabricated landscapes); and in Ingham et al., 2007, Proc. Natl. Acad. Sci. USA 104: 18217-18222 (the micro-Petri dish, a million-well growth chip for the culture and high-throughput screening of microorganisms).